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Although large deletions in the dystrophin gene have been identified in more than two-thirds of Duchenne and Becker muscular dystrophy patients, the molecular mechanisms that lead to the generation of these deletions are largely unknown. Here, Alu and LINE-1 (L1) repetitive elements were shown to be present at one or other of the two ends, respectively, of a 430-kb deletion in the dystrophin gene. The breakpoint of the deletion, which stretches from exons 2 to 7, was defined more precisely by polymerase chain reaction (PCR) walking on introns 1 and 7. Finally, the region containing the breakpoint was amplified as a fragment of more than 10 kb. Sequencing of the deletion endpoint revealed the presence of an Alu sequence in intron 1, 25 kb downstream from the 3Ј end of exon 1 that was joined directly to an L1 sequence in intron 7, 4.5 kb downstream from the 3Ј end of exon 7. The deletion was calculated to be 430 kb. To our knowledge, this is a novel recombination event joining non-homologous Alu and L1 repeats, and is the largest known intrachromosomal deletion that is thought to involve repetitive genetic elements. Sequence characteristics around the breakpoint are discussed.

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To date, many mutations, including intronic nucleotide changes, in the SLC12A3 gene encoding the thiazide-sensitive sodium-chloride cotransporter (NCCT) have been reported in Gitelman's syndrome (GS) patients. However, it has not been clarified whether intronic nucleotide changes affect mRNA content. Since mRNA analysis is possible only after obtaining renal biopsy specimens, no studies have been conducted to identify transcript abnormalities in GS. In the study reported here, we investigated such transcript abnormalities for the first time by using mRNA expressed in a patient's urinary sediment cells. Direct sequencing analysis of leukocyte DNA disclosed one known missense mutation (R399C) and one known nucleotide change of the splicing acceptor site of intron 13 (1670-1 g > t). mRNA extracted from the urinary sediment cells was analyzed by RT-PCR to determine the pathogenic role of the intron mutation. A fragment encompassing exon 13 to 15 was amplified as two products, one consisting of all three exons and the other lacking only exon 14 in its entirety. Our investigation was the first to demonstrate exon 14 skipping in an NCCT transcript in renal cells. This methodology thus constitutes a potential noninvasive analytical tool for every inherited kidney disease.